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Substantial decreases in the number and diversity of microbiota during chemotherapy-induced gastrointestinal mucositis in a rat model

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Abstract

Purpose

Earlier, we showed in acute myeloid leukemia (AML) patients that the microbiota changes dramatically during anticancer treatment, coinciding with gastrointestinal mucositis: The commensal anaerobic populations reduce in favor of potential pathogens. Therefore, interventions targeting the microbiota during mucositis might be interesting but can better be tested in animals than in vulnerable mucositis patients. Here, we aimed to study the potential microbial changes during methotrexate (MTX)-induced gastrointestinal mucositis in a well-established rat model and to study whether this model can be used for future microbial intervention studies.

Methods

After injection with MTX or saline (day 0), rats were sacrificed between days 2 and 11. Plasma citrulline level, jejunal histology, and the number and diversity of intestinal bacteria in feces (using fluorescence in situ hybridization (FISH)) were determined.

Results

Mucositis was most severe on day 4 when food intake, plasma citrulline, and villus length were the lowest, compared with controls (P < 0.0125). At the same time, MTX-treated rats showed an overall decrease (705-fold) in most bacteria (using a universal probe), compared with controls (P < 0.125). Reduced bacterial presence was related with the presence of diarrhea and a reduced villus length (rho = 0.38, P < 0.05). At day 4, there was an absolute and relative decrease of anaerobes (13-fold and −58 %, respectively) and streptococci (296-fold and −1 %, respectively) but a relative increase of Bacteroides (+49 %), compared with controls (P < 0.125).

Conclusions

In the mucositis rat model, we found substantial decreases in the number and diversity of microbiota, resembling earlier findings in humans. The model therefore seems well suited to study the effects of different microbial interventions on mucositis, prior to performing human studies.

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Abbreviations

FISH:

Fluorescence in situ hybridization

H&E:

Hematoxylin and eosin

MTX:

Methotrexate

n :

Number of rats per group

SPF:

Specific-pathogen-free

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Acknowledgments

The authors wish to thank Rick Havinga, Juul Baller, and Pieter Klok (University Medical Center Groningen, the Netherlands) for excellent technical assistance in our studies.

Conflict of interest

This work was financially supported by an unrestricted research grant from Fonds NutsOhra (the Netherlands). We confirm that no one from Fonds NutsOhra was involved in the planning of the study, the collection or analysis of study data, or the preparation of the manuscript. We have nothing to declare.

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Authors and Affiliations

  1. Department of Pediatrics, Beatrix Children’s Hospital, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands

    Margot Fijlstra & Wim J. E. Tissing

  2. Department of Pediatrics, Emma Children’s Hospital, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands

    Margot Fijlstra

  3. Department of Pediatrics, Sophia Children’s Hospital, Erasmus Medical Center, University of Rotterdam, Rotterdam, The Netherlands

    Edmond H. H. M. Rings

  4. Department of Pediatrics, Willem-Alexander Children’s Hospital, Leiden University Medical Center, Leiden University, Leiden, The Netherlands

    Edmond H. H. M. Rings

  5. Department of Medical Microbiology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands

    Mithila Ferdous, Anne M. Koning & Hermie J. M. Harmsen

  6. Beatrix Children’s Hospital, University Medical Center Groningen, HPC CA21, P.O. Box 30.001, 9700 RB, Groningen, The Netherlands

    Wim J. E. Tissing

Authors
  1. Margot Fijlstra
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  2. Mithila Ferdous
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  4. Edmond H. H. M. Rings
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Correspondence to Wim J. E. Tissing.

Additional information

Relevance of our manuscript to inform research, policies, and/or programs

In this manuscript, we demonstrate that rats with gastrointestinal mucositis show substantial decreases in the number and diversity of microbiota, thereby favoring potentially enteropathogenic bacteria. Our findings resemble earlier findings in mucositis patients. The model therefore seems well suited to study the microbiota in relation to mucositis and to study if microbial interventions (like the use of prebiotics, probiotics, and antibiotics) can change the course of mucositis, prior to performing selected studies in vulnerable mucositis patients. Our work is relevant for everyone who is involved in the caretaking of patients with mucositis (doctors, dieticians, other caretakers) but especially for producers of agents aiming at restoring dysbioisis and/or enhancing recovery from mucositis (food industries, pharmacologists, researchers, etc.): The effects of their products on mucositis might well be studied in the mucositis model.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Online Resource 1

Clinical and histological findings in the mucositis rat model. Food intake (A), body weight (relative to weight at day 0, B), plasma citrulline concentration (C) and jejunal crypt and villus length (D and E respectively) in saline-treated rats (controls, ○ ---, n = 12) and methotrexate-treated rats (MTX, ● ▬, n = 21), up to 10 days after i.v. saline- or MTX-injection (day 0, see ‘Materials and methods’). Intake is shown until day 9, since rats were terminated at day 10 or 11. Data represent medians and first to third quartiles (p25-p75, A-B) or medians and data for individual rats (C-E). # P < 0.05 for controls versus MTX-treated rats, *P < 0.0125 for controls versus MTX-treated rats. (PDF 58 kb)

Online Resource 2

Correlation between the absolute number of most bacteria and villus length in the mucositis rat model. Data correspond with data from Online Resource 1E (villus length) and Fig. 1a (most bacteria, using a universal probe), in saline-treated rats (controls, ○, n = 12), methotrexate-treated rats with diarrhea (MTX, ■, n = 5) and methotrexate-treated rats without diarrhea (MTX, ●, n = 16). The Spearman-correlation (r) and P-value is indicated. (PDF 36 kb)

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Fijlstra, M., Ferdous, M., Koning, A.M. et al. Substantial decreases in the number and diversity of microbiota during chemotherapy-induced gastrointestinal mucositis in a rat model. Support Care Cancer 23, 1513–1522 (2015). https://doi.org/10.1007/s00520-014-2487-6

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